Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 7.132
Filtrar
1.
Nature ; 579(7798): 291-296, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32103174

RESUMO

The DNA-dependent protein kinase (DNA-PK), which comprises the KU heterodimer and a catalytic subunit (DNA-PKcs), is a classical non-homologous end-joining (cNHEJ) factor1. KU binds to DNA ends, initiates cNHEJ, and recruits and activates DNA-PKcs. KU also binds to RNA, but the relevance of this interaction in mammals is unclear. Here we use mouse models to show that DNA-PK has an unexpected role in the biogenesis of ribosomal RNA (rRNA) and in haematopoiesis. The expression of kinase-dead DNA-PKcs abrogates cNHEJ2. However, most mice that both expressed kinase-dead DNA-PKcs and lacked the tumour suppressor TP53 developed myeloid disease, whereas all other previously characterized mice deficient in both cNHEJ and TP53 expression succumbed to pro-B cell lymphoma3. DNA-PK autophosphorylates DNA-PKcs, which is its best characterized substrate. Blocking the phosphorylation of DNA-PKcs at the T2609 cluster, but not the S2056 cluster, led to KU-dependent defects in 18S rRNA processing, compromised global protein synthesis in haematopoietic cells and caused bone marrow failure in mice. KU drives the assembly of DNA-PKcs on a wide range of cellular RNAs, including the U3 small nucleolar RNA, which is essential for processing of 18S rRNA4. U3 activates purified DNA-PK and triggers phosphorylation of DNA-PKcs at T2609. DNA-PK, but not other cNHEJ factors, resides in nucleoli in an rRNA-dependent manner and is co-purified with the small subunit processome. Together our data show that DNA-PK has RNA-dependent, cNHEJ-independent functions during ribosome biogenesis that require the kinase activity of DNA-PKcs and its phosphorylation at the T2609 cluster.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Hematopoese/genética , Autoantígeno Ku/metabolismo , Linfoma/enzimologia , Linfoma/fisiopatologia , RNA Ribossômico 18S/metabolismo , Proteínas de Ligação ao Cálcio/genética , Domínio Catalítico/fisiologia , Reparo do DNA/genética , Ativação Enzimática/genética , Células HeLa , Humanos , Linfoma/genética , Modelos Animais , Mutação , Fosforilação , Ligação Proteica , Biossíntese de Proteínas/genética , RNA Ribossômico 18S/genética , RNA Nucleolar Pequeno/metabolismo
2.
Chem Biol Interact ; 318: 108977, 2020 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-32035863

RESUMO

Excess weight and obesity increase the risk of developing major risk factors for chronic kidney disease. Lignin comprises 20%-30% of the global plant biomass; however, it is not well utilized because of its resistance to chemical and biological degradation. We investigated whether low-molecular-weight oxidized lignophenol (LOLP), a lignin derivative, could alter inflammation and fibrosis in the kidneys of a high-fat diet (HFD)-fed mice. Male mice were divided into three treatment groups: HFD; HFD +0.3% LOLP; and HFD +0.6% LOLP. The control mice (Cont) were fed a low-fat diet. Macrophage kinetics, the degree of fibrosis, the extent of phosphorylation of AMP-activated protein kinase (AMPK), and mRNA expression of proinflammatory mediators in the kidneys were examined. The number of macrophages, the percentage of fibrotic area, and the mRNA expression of proinflammatory markers, TNF-α and Ccl2, and a marker of fibrosis, TGF-ß, were significantly higher in the kidneys of mice in the HFD group than those in the Cont group. Conversely, treatment with 0.6% LOLP for 8 weeks significantly suppressed the degree of macrophage infiltration, interstitial fibrotic area, and the increased mRNA expression of proinflammatory and fibrosis markers induced by HFD. In conclusion, LOLP suppressed macrophage infiltration and the increase in fibrotic area, and upregulated AMPK phosphorylation in the kidneys of HFD-fed mice; thus, it may ameliorate HFD-induced kidney injury.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Dieta Hiperlipídica/efeitos adversos , Nefropatias/induzido quimicamente , Lignina/química , Fenóis/química , Fenóis/farmacologia , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Ativação Enzimática/efeitos dos fármacos , Fibrose/induzido quimicamente , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Nefropatias/prevenção & controle , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro , Receptores Acoplados a Proteínas-G/genética , Receptores Acoplados a Proteínas-G/metabolismo
3.
Virchows Arch ; 476(1): 17-28, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31463731

RESUMO

The classification of vascular neoplasms continues to evolve as we accumulate more genetic and clinical data, particularly for rare tumor types. Because of tumor rarity, changes to classification schema, overlapping histologic features, and in some cases, lack of morphologic evidence of vasoformation, vascular neoplasms present a diagnostic challenge. Here, we discuss recent developments in our understanding of vascular tumors, with a detailed discussion of epithelioid hemangioma, tufted angioma, kaposiform hemangioendothelioma, composite hemangioendothelioma, pseudomyogenic hemangioendothelioma, epithelioid hemangioendothelioma, and angiosarcoma.


Assuntos
Neoplasias Vasculares/patologia , Proteínas de Ligação ao Cálcio/genética , Hemangioendotelioma/genética , Hemangioendotelioma/patologia , Hemangioendotelioma Epitelioide/genética , Hemangioendotelioma Epitelioide/patologia , Hemangioma/genética , Hemangioma/patologia , Hemangiossarcoma/genética , Hemangiossarcoma/patologia , Humanos , Síndrome de Kasabach-Merritt/genética , Síndrome de Kasabach-Merritt/patologia , Proteínas Proto-Oncogênicas c-myc/análise , Sarcoma de Kaposi/genética , Sarcoma de Kaposi/patologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Transativadores/genética , Neoplasias Vasculares/genética
4.
Gene ; 723: 144120, 2020 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-31589964

RESUMO

PURPOSE: Matrix Gla protein (MGP) is a vitamin K-dependent, γ-carboxylated protein that was initially found to be a physiological inhibitor of ectopic calcifications affecting mainly cartilage and the vascular system. Mutations in the MGP gene were found to be responsible for a human pathology, the Keutel syndrome, characterized by abnormal calcifications in cartilage, lungs, brain and vascular system. MGP was recently implicated in tumorigenic processes such as angiogenesis and shown to be abnormally regulated in several tumors, including cervical, ovarian, urogenital and breast. This fact has triggered our interest in analyzing the expression of MGP and of its regulator, the transcription factor runt related transcription factor 2 (RUNX2), in colorectal cancer (CRC). METHODS: MGP and RUNX2 expression were analyzed in cancer and non-tumor biopsies samples from 33 CRC patients and 9 healthy controls by RT-qPCR. Consequently, statistical analyses were performed to evaluate the clinical-pathological significance of MGP and RUNX2 in CRC. MGP protein was also detected by immunohistochemical analysis. RESULTS: Showed an overall overexpression of MGP in the tumor mucosa of patients at mRNA level when compared to adjacent normal mucosa and healthy control tissues. In addition, analysis of the expression of RUNX2 mRNA demonstrated an overexpression in CRC tissue samples and a positive correlation with MGP expression (Pearson correlation coefficient 0.636; p ≤ 0.01) in tumor mucosa. However correlations between MGP gene expression and clinical-pathological characteristics, such as gender, age and pathology classification did not provide relevant information that may shed light towards the differences of MGP expression observed between normal and malignant tissue. CONCLUSIONS: We were able to associate the high levels of MGP mRNA expression with a worse prognosis and survival rate lower than five years. These results contributed to improve our understanding of the molecular mechanism underlying MGP deregulation in cancer.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Neoplasias Colorretais/patologia , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Regulação para Cima , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Análise de Sobrevida
5.
Nat Genet ; 51(12): 1679-1690, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31784728

RESUMO

NRXN1 undergoes extensive alternative splicing, and non-recurrent heterozygous deletions in NRXN1 are strongly associated with neuropsychiatric disorders. We establish that human induced pluripotent stem cell (hiPSC)-derived neurons well represent the diversity of NRXN1α alternative splicing observed in the human brain, cataloguing 123 high-confidence in-frame human NRXN1α isoforms. Patient-derived NRXN1+/- hiPSC-neurons show a greater than twofold reduction in half of the wild-type NRXN1α isoforms and express dozens of novel isoforms from the mutant allele. Reduced neuronal activity in patient-derived NRXN1+/- hiPSC-neurons is ameliorated by overexpression of individual control isoforms in a genotype-dependent manner, whereas individual mutant isoforms decrease neuronal activity levels in control hiPSC-neurons. In a genotype-dependent manner, the phenotypic impact of patient-specific NRXN1+/- mutations can occur through a reduction in wild-type NRXN1α isoform levels as well as the presence of mutant NRXN1α isoforms.


Assuntos
Processamento Alternativo , Proteínas de Ligação ao Cálcio/genética , Células-Tronco Pluripotentes Induzidas/fisiologia , Moléculas de Adesão de Célula Nervosa/genética , Esquizofrenia/genética , Animais , Transtorno do Espectro Autista/genética , Transtorno Bipolar/genética , Estudos de Casos e Controles , Transtorno Depressivo Maior/genética , Feminino , Expressão Gênica , Heterozigoto , Humanos , Masculino , Camundongos , Isoformas de Proteínas/genética , Deleção de Sequência
6.
BMC Med Genet ; 20(1): 188, 2019 11 27.
Artigo em Inglês | MEDLINE | ID: mdl-31775791

RESUMO

BACKGROUND: Gorlin-Chaudhry-Moss syndrome (GCMS) and Fontaine-Farriaux syndrome (FFS) are extremely rare genetic disorders that share similar clinical manifestations. Because a de novo missense mutation of the solute carrier family 25 member 24 (SLC25A24) gene was suggested to be the common genetic basis of both syndromes, it has been proposed recently that they be integrated into a single disorder under the name of Fontaine progeroid syndrome (FPS). CASE PRESENTATION: A 9-year-old Korean girl presented with typical clinical features of FPS. She had generalized loose skin with decreased subcutaneous fat, skin wrinkling on the forehead and limbs, skull deformities and a peculiar facial appearance with microphthalmia and midface hypoplasia, anomalies of the digits and nails, a large umbilical hernia and a nearly normal developmental outcome. She exhibited prenatal and postnatal growth retardation together with short stature, and records showed that her height and weight were invariably under - 2.0 SD from birth to the age of 10 years. SLC25A24 analysis revealed a heterozygous mutation reported previously, NM_013386:c.650G > A, p.[Arg217His]. After screening her family for the identified mutation, she was confirmed as being a de novo case of FPS caused by an SLC25A24 mutation. CONCLUSION: We describe a Korean girl with typical clinical findings of FPS and a de novo mutation in SLC25A24, as well as 10 years of clinical follow-up, including growth and developmental achievements.


Assuntos
Antiporters/genética , Proteínas de Ligação ao Cálcio/genética , Proteínas Mitocondriais/genética , Fenótipo , Progéria/patologia , Criança , Feminino , Seguimentos , Humanos , Masculino , Mutação de Sentido Incorreto , Linhagem , Progéria/genética , República da Coreia , Síndrome
7.
PLoS Biol ; 17(10): e3000466, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31658245

RESUMO

The pre- and postsynaptic membranes comprising the synaptic junction differ in protein composition. The membrane trafficking mechanisms by which neurons control surface polarization of synaptic receptors remain poorly understood. The sorting receptor Sortilin-related CNS expressed 1 (SorCS1) is a critical regulator of trafficking of neuronal receptors, including the presynaptic adhesion molecule neurexin (Nrxn), an essential synaptic organizer. Here, we show that SorCS1 maintains a balance between axonal and dendritic Nrxn surface levels in the same neuron. Newly synthesized Nrxn1α traffics to the dendritic surface, where it is endocytosed. Endosomal SorCS1 interacts with the Rab11 GTPase effector Rab11 family-interacting protein 5 (Rab11FIP5)/Rab11 interacting protein (Rip11) to facilitate the transition of internalized Nrxn1α from early to recycling endosomes and bias Nrxn1α surface polarization towards the axon. In the absence of SorCS1, Nrxn1α accumulates in early endosomes and mispolarizes to the dendritic surface, impairing presynaptic differentiation and function. Thus, SorCS1-mediated sorting in dendritic endosomes controls Nrxn axonal surface polarization required for proper synapse development and function.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Córtex Cerebral/metabolismo , Moléculas de Adesão de Célula Nervosa/genética , Neurônios/metabolismo , Receptores de Superfície Celular/genética , Membranas Sinápticas/metabolismo , Transmissão Sináptica/genética , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Polaridade Celular , Córtex Cerebral/citologia , Embrião de Mamíferos , Endocitose , Endossomos/metabolismo , Regulação da Expressão Gênica , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Moléculas de Adesão de Célula Nervosa/metabolismo , Neurônios/ultraestrutura , Cultura Primária de Células , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transporte Proteico , Ratos , Ratos Wistar , Receptores de Superfície Celular/metabolismo , Membranas Sinápticas/ultraestrutura , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
8.
EMBO J ; 38(22): e101603, 2019 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-31566781

RESUMO

Neurexins are presynaptic, cell-adhesion molecules that specify the functional properties of synapses via interactions with trans-synaptic ligands. Neurexins are extensively alternatively spliced at six canonical sites that regulate multifarious ligand interactions, but the structural mechanisms underlying alternative splicing-dependent neurexin regulation are largely unknown. Here, we determined high-resolution structures of the complex of neurexophilin-1 and the second laminin/neurexin/sex-hormone-binding globulin domain (LNS2) of neurexin-1 and examined how alternative splicing at splice site #2 (SS2) regulates the complex. Our data reveal a unique, extensive, neurexophilin-neurexin binding interface that extends the jelly-roll ß-sandwich of LNS2 of neurexin-1 into neurexophilin-1. The SS2A insert of LNS2 augments this interface, increasing the binding affinity of LNS2 for neurexophilin-1. Taken together, our data reveal an unexpected architecture of neurexophilin-neurexin complexes that accounts for the modulation of binding by alternative splicing, which in turn regulates the competition of neurexophilin for neurexin binding with other ligands.


Assuntos
Processamento Alternativo , Proteínas de Ligação ao Cálcio/química , Proteínas de Ligação ao Cálcio/metabolismo , Glicoproteínas/química , Glicoproteínas/metabolismo , Laminina/metabolismo , Moléculas de Adesão de Célula Nervosa/química , Moléculas de Adesão de Célula Nervosa/metabolismo , Neuropeptídeos/química , Neuropeptídeos/metabolismo , Sequência de Aminoácidos , Animais , Proteínas de Ligação ao Cálcio/genética , Cristalografia por Raios X , Glicoproteínas/genética , Ligantes , Camundongos , Modelos Moleculares , Moléculas de Adesão de Célula Nervosa/genética , Neuropeptídeos/genética , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Ratos , Homologia de Sequência
9.
Nat Commun ; 10(1): 4910, 2019 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-31659163

RESUMO

AspH is an endoplasmic reticulum (ER) membrane-anchored 2-oxoglutarate oxygenase whose C-terminal oxygenase and tetratricopeptide repeat (TPR) domains present in the ER lumen. AspH catalyses hydroxylation of asparaginyl- and aspartyl-residues in epidermal growth factor-like domains (EGFDs). Here we report crystal structures of human AspH, with and without substrate, that reveal substantial conformational changes of the oxygenase and TPR domains during substrate binding. Fe(II)-binding by AspH is unusual, employing only two Fe(II)-binding ligands (His679/His725). Most EGFD structures adopt an established fold with a conserved Cys1-3, 2-4, 5-6 disulfide bonding pattern; an unexpected Cys3-4 disulfide bonding pattern is observed in AspH-EGFD substrate complexes, the catalytic relevance of which is supported by studies involving stable cyclic peptide substrate analogues and by effects of Ca(II) ions on activity. The results have implications for EGFD disulfide pattern processing in the ER and will enable medicinal chemistry efforts targeting human 2OG oxygenases.


Assuntos
Proteínas de Ligação ao Cálcio/química , Proteínas de Membrana/química , Oxigenases de Função Mista/química , Proteínas Musculares/química , Sequência de Aminoácidos , Asparagina/metabolismo , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Domínio Catalítico , Cristalografia , Dissulfetos/química , Dissulfetos/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Compostos Ferrosos/química , Compostos Ferrosos/metabolismo , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Conformação Proteica
10.
Nucleic Acids Res ; 47(19): 10151-10165, 2019 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-31665741

RESUMO

RAD51 plays a central role in homologous recombination during double-strand break repair and in replication fork dynamics. Misregulation of RAD51 is associated with genetic instability and cancer. RAD51 is regulated by many accessory proteins including the highly conserved Shu complex. Here, we report the function of the human Shu complex during replication to regulate RAD51 recruitment to DNA repair foci and, secondly, during replication fork restart following replication fork stalling. Deletion of the Shu complex members, SWS1 and SWSAP1, using CRISPR/Cas9, renders cells specifically sensitive to the replication fork stalling and collapse caused by methyl methanesulfonate and mitomycin C exposure, a delayed and reduced RAD51 response, and fewer sister chromatid exchanges. Our additional analysis identified SPIDR and PDS5B as novel Shu complex interacting partners and genetically function in the same pathway upon DNA damage. Collectively, our study uncovers a protein complex, which consists of SWS1, SWSAP1, SPIDR and PDS5B, involved in DNA repair and provides insight into Shu complex function and composition.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação a DNA/genética , Recombinação Homóloga/genética , Proteínas Nucleares/genética , Recombinases Rec A/genética , Fatores de Transcrição/genética , Sistemas CRISPR-Cas/genética , Dano ao DNA/genética , Reparo do DNA/genética , Replicação do DNA/genética , Instabilidade Genômica/genética , Humanos , Complexos Multiproteicos/genética , Rad51 Recombinase/genética , Troca de Cromátide Irmã/genética
11.
Int J Mol Sci ; 20(19)2019 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-31569565

RESUMO

Fifteen transcription factors in the CAMTA (calmodulin binding transcription activator) family of soybean were reported to differentially regulate in multiple stresses; however, their functional analyses had not yet been attempted. To characterize their role in stresses, we first comprehensively analyzed the GmCAMTA family in silico and thereafter determined their expression pattern under drought. The bioinformatics analysis revealed multiple stress-related cis-regulatory elements including ABRE, SARE, G-box and W-box, 10 unique miRNA (microRNA) targets in GmCAMTA transcripts and 48 proteins in GmCAMTAs' interaction network. We then cloned the 2769 bp CDS (coding sequence) of GmCAMTA12 in an expression vector and overexpressed in soybean and Arabidopsis through Agrobacterium-mediated transformation. The T3 (Transgenic generation 3) stably transformed homozygous lines of Arabidopsis exhibited enhanced tolerance to drought in soil as well as on MS (Murashige and Skoog) media containing mannitol. In their drought assay, the average survival rate of transgenic Arabidopsis lines OE5 and OE12 (Overexpression Line 5 and Line 12) was 83.66% and 87.87%, respectively, which was ~30% higher than that of wild type. In addition, the germination and root length assays as well as physiological indexes such as proline and malondialdehyde contents, catalase activity and leakage of electrolytes affirmed the better performance of OE lines. Similarly, GmCAMTA12 overexpression in soybean promoted drought-efficient hairy roots in OE chimeric plants as compare to that of VC (Vector control). In parallel, the improved growth performance of OE in Hoagland-PEG (polyethylene glycol) and on MS-mannitol was revealed by their phenotypic, physiological and molecular measures. Furthermore, with the overexpression of GmCAMTA12, the downstream genes including AtAnnexin5, AtCaMHSP, At2G433110 and AtWRKY14 were upregulated in Arabidopsis. Likewise, in soybean hairy roots, GmELO, GmNAB and GmPLA1-IId were significantly upregulated as a result of GmCAMTA12 overexpression and majority of these upregulated genes in both plants possess CAMTA binding CGCG/CGTG motif in their promoters. Taken together, we report that GmCAMTA12 plays substantial role in tolerance of soybean against drought stress and could prove to be a novel candidate for engineering soybean and other plants against drought stress. Some research gaps were also identified for future studies to extend our comprehension of Ca-CaM-CAMTA-mediated stress regulatory mechanisms.


Assuntos
Adaptação Biológica/genética , Arabidopsis/fisiologia , Proteínas de Ligação ao Cálcio/genética , Secas , Expressão Gênica , Soja/fisiologia , Estresse Fisiológico/genética , Sequência de Aminoácidos , Arabidopsis/classificação , Proteínas de Ligação ao Cálcio/química , Proteínas de Ligação ao Cálcio/metabolismo , Fenômenos Químicos , Filogenia , Soja/classificação
12.
J Agric Food Chem ; 67(40): 11035-11043, 2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31517486

RESUMO

Ca2+-binding proteins (CaBPs) are widely distributed as Ca2+ sensor relay proteins that regulate various cellular processes, including Ca2+ homeostasis. Diamide insecticides such as cyantraniliprole kill insects by disrupting the Ca2+ homeostasis in muscle cells. However, less attention has been paid to the roles of CaBPs in response to insecticides. In this study, two CaBP genes (BtCaBP1 and BtCaBP2) were identified in the whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae), and their functions in response to cyantraniliprole were investigated. After expression of BtCaBP1 and BtCaBP2 in vitro, the results of Ca2+ imaging and cytotoxicity assay revealed that the overexpression of each of the BtCaBPs stabilized Ca2+ concentration in the cytoplasm after exposure to cyantraniliprole and decreased the toxicity of cyantraniliprole against Sf9 cells. However, the knockdown of BtCaBP1 or BtCaBP2 in vivo significantly increased the toxicity of cyantraniliprole to B. tabaci. Taken together, these results provide evidence that BtCaBP1 and BtCaBP2 play a role in response to cyantraniliprole exposure through stabilization of Ca2+ concentration in whiteflies.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Clonagem Molecular , Hemípteros/efeitos dos fármacos , Hemípteros/genética , Proteínas de Insetos/metabolismo , Inseticidas/farmacologia , Pirazóis/farmacologia , ortoaminobenzoatos/farmacologia , Sequência de Aminoácidos , Animais , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/genética , Hemípteros/classificação , Hemípteros/metabolismo , Proteínas de Insetos/genética , Filogenia
14.
Nat Chem Biol ; 15(10): 992-1000, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31527837

RESUMO

Post-translational modifications of histone variant H2A.Z accompany gene transactivation, but its modifying enzymes still remain elusive. Here, we reveal a hitherto unknown function of human KAT2A (GCN5) as a histone acetyltransferase (HAT) of H2A.Z at the promoters of a set of transactivated genes. Expression of these genes also depends on the DNA repair complex XPC-RAD23-CEN2. We established that XPC-RAD23-CEN2 interacts both with H2A.Z and KAT2A to drive the recruitment of the HAT at promoters and license H2A.Z acetylation. KAT2A selectively acetylates H2A.Z.1 versus H2A.Z.2 in vitro on several well-defined lysines and we unveiled that alanine-14 in H2A.Z.2 is responsible for inhibiting the activity of KAT2A. Notably, the use of a nonacetylable H2A.Z.1 mutant shows that H2A.Z.1ac recruits the epigenetic reader BRD2 to promote RNA polymerase II recruitment. Our studies identify KAT2A as an H2A.Z.1 HAT in mammals and implicate XPC-RAD23-CEN2 as a transcriptional co-activator licensing the reshaping of the promoter epigenetic landscape.


Assuntos
Reparo do DNA/fisiologia , Histona Acetiltransferases/metabolismo , Histonas/metabolismo , Acetilação , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Enzimas Reparadoras do DNA/genética , Enzimas Reparadoras do DNA/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Fibroblastos , Regulação da Expressão Gênica , Humanos , Lisina Acetiltransferase 5
15.
BMC Cancer ; 19(1): 898, 2019 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-31500591

RESUMO

BACKGROUND: Although the leucine zipper-EF-hand-containing transmembrane protein 1 (LETM1) is one of the mitochondrial inner membrane proteins that is involved in cancer prognosis in various tumors, LETM1 as a biomarker for prognostic evaluation of non-small cell lung carcinoma (NSCLC) has not been well studied. METHODS: To address this issue, we used 75 cases NSCLC, 20 cases adjacent normal lung tissues and NSCLC cell lines. We performed immunohistochemistry staining and western blot analysis as well as immunofluorescence imaging. RESULTS: Our studies show that expression of LETM1 is significantly correlated with the lymph node metastasis (p = 0.003) and the clinical stage (p = 0.005) of NSCLC. The Kaplan-Meier survival analysis revealed that NSCLC patients with positive expression of LETM1 exhibits a shorter overall survival (OS) rate (p = 0.005). The univariate and multivariate Cox regression analysis indicated that LETM1 is a independent poor prognostic marker of NSCLC. In addition, the LETM1 expression is correlated with cancer stemness-related gene LGR5 (p < 0.001) and HIF1α expression (p < 0.001), but not with others. Moreover, LETM1 expression was associated with the expression of cyclin D1 (p = 0.003), p27 (p = 0.001), pPI3K(p85) (p = 0.025), and pAkt-Thr308 (p = 0.004). Further, our studies show in LETM1-positive NSCLC tissues the microvessel density was significantly higher than in the negative ones (p = 0.024). CONCLUSION: These results indicate that LETM1 is a potential prognostic biomarker of NSCLC.


Assuntos
Proteínas de Ligação ao Cálcio/análise , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas de Membrana/análise , Idoso , Biomarcadores Tumorais/análise , Proteínas de Ligação ao Cálcio/genética , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/análise , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Prognóstico , Receptores Acoplados a Proteínas-G/análise , Receptores Acoplados a Proteínas-G/genética
16.
Cell Tissue Res ; 378(2): 267-277, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31392520

RESUMO

Peeling skin syndrome is a heterogeneous group of rare disorders. Peeling skin, leukonychia, acral punctate keratoses, cheilitis and knuckle pads (PLACK syndrome, OMIM616295) is a newly described form of PSS with an autosomal recessive mode of inheritance. We report a 5.5-year-old boy with features of PLACK syndrome. Additionally, he had mild cerebral atrophy and mild muscle involvements. Whole exome sequencing was performed in genomic DNA of this individual and subsequent analysis revealed a homozygous c.544G > T (p.Glu182*) nonsense mutation in the CAST gene encoding calpastatin. Sanger sequencing confirmed this variant and demonstrated that his affected aunt was also homozygous. Real-time qRT-PCR and immunoblot analysis showed reduced calpastatin expression in skin fibroblasts derived from both affected individuals compared to heterozygous family members. In vitro calpastatin activity assays also showed decreased activity in affected individuals. This study further supports a key role for calpastatin in the tight regulation of proteolytic pathways within the skin.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Códon sem Sentido/genética , Dermatite Esfoliativa/genética , Dermatopatias Genéticas/genética , Pele , Adulto , Pré-Escolar , Feminino , Homozigoto , Humanos , Masculino , Pele/metabolismo , Pele/patologia
17.
Science ; 365(6455): 786-793, 2019 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-31395745

RESUMO

How cellular and organismal complexity emerges from combinatorial expression of genes is a central question in biology. High-content phenotyping approaches such as Perturb-seq (single-cell RNA-sequencing pooled CRISPR screens) present an opportunity for exploring such genetic interactions (GIs) at scale. Here, we present an analytical framework for interpreting high-dimensional landscapes of cell states (manifolds) constructed from transcriptional phenotypes. We applied this approach to Perturb-seq profiling of strong GIs mined from a growth-based, gain-of-function GI map. Exploration of this manifold enabled ordering of regulatory pathways, principled classification of GIs (e.g., identifying suppressors), and mechanistic elucidation of synergistic interactions, including an unexpected synergy between CBL and CNN1 driving erythroid differentiation. Finally, we applied recommender system machine learning to predict interactions, facilitating exploration of vastly larger GI manifolds.


Assuntos
Epistasia Genética , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Apoptose/genética , Sistemas CRISPR-Cas , Proteínas de Ligação ao Cálcio/genética , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Células Eritroides/citologia , Eritropoese/genética , Feminino , Perfilação da Expressão Gênica , Granulócitos/citologia , Humanos , Proteínas dos Microfilamentos/genética , Proteínas Proto-Oncogênicas c-cbl/genética
18.
Int J Mol Sci ; 20(15)2019 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-31370244

RESUMO

Aneurysmal subarachnoid hemorrhage (aSAH), characterized by the extravasation of blood into the subarachnoid space caused by an intracranial aneurysm rupture, may lead to neurocognitive impairments and permanent disability and usually carries poor outcome. Dental or gingiva-derived stem cells have been shown to contribute to immune modulation and neuroregeneration, but the underlying mechanisms are unclear. In the present study, we sought to investigate whether dental pulp stem cells (DPSCs) secrete certain factor(s) that can ameliorate the neural damage and other manifestations in a rat aSAH model. Twenty-four hours after the induction of aSAH, microthrombosis, cortical vasoconstriction, and the decrease in microcirculation and tissue oxygen pressure were detected. Intrathecal administration of DPSC-derived conditioned media (DPSC-CM) ameliorated aSAH-induced vasoconstriction, neuroinflammation, and improved the oxygenation in the injured brain. Rotarod test revealed that the aSAH-induced cognitive and motor impairments were significantly improved by this DPSC-CM administration. Cytokine array indicated the major constituent of DPSC-CM was predominantly insulin growth factor-1 (IGF-1). Immunohistochemistry staining of injured brain tissue revealed the robust increase in Iba1-positive cells that were also ameliorated by DPSC-CM administration. Antibody-mediated neutralization of IGF-1 moderately deteriorated the rescuing effect of DPSC-CM on microcirculation, Iba1-positive cells in the injured brain area, and the cognitive/motor impairments. Taken together, the DPSC-derived secretory factors showed prominent therapeutic potential for aSAH. This therapeutic efficacy may include improvement of microcirculation, alleviation of neuroinflammation, and microglial activation; partially through IGF-1-dependent mechanisms.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Meios de Cultivo Condicionados/farmacologia , Transtornos Neurocognitivos/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Transtornos Psicomotores/tratamento farmacológico , Hemorragia Subaracnóidea/tratamento farmacológico , Trombose/tratamento farmacológico , Animais , Isquemia Encefálica/genética , Isquemia Encefálica/metabolismo , Isquemia Encefálica/fisiopatologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Meios de Cultivo Condicionados/química , Polpa Dentária/citologia , Polpa Dentária/metabolismo , Modelos Animais de Doenças , Expressão Gênica , Injeções Espinhais , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Microcirculação/efeitos dos fármacos , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Transtornos Neurocognitivos/genética , Transtornos Neurocognitivos/metabolismo , Transtornos Neurocognitivos/fisiopatologia , Fármacos Neuroprotetores/química , Consumo de Oxigênio/efeitos dos fármacos , Transtornos Psicomotores/genética , Transtornos Psicomotores/metabolismo , Transtornos Psicomotores/fisiopatologia , Ratos , Ratos Wistar , Teste de Desempenho do Rota-Rod , Células-Tronco/química , Células-Tronco/citologia , Células-Tronco/metabolismo , Hemorragia Subaracnóidea/genética , Hemorragia Subaracnóidea/metabolismo , Hemorragia Subaracnóidea/fisiopatologia , Trombose/genética , Trombose/metabolismo , Trombose/fisiopatologia , Vasoconstrição/efeitos dos fármacos
19.
Nat Commun ; 10(1): 3596, 2019 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399601

RESUMO

Stem cells (SCs) receive inductive cues from the surrounding microenvironment and cells. Limited molecular evidence has connected tissue-specific mesenchymal stem cells (MSCs) with mesenchymal transit amplifying cells (MTACs). Using mouse incisor as the model, we discover a population of MSCs neibouring to the MTACs and epithelial SCs. With Notch signaling as the key regulator, we disclose molecular proof and lineage tracing evidence showing the distinct MSCs contribute to incisor MTACs and the other mesenchymal cell lineages. MTACs can feedback and regulate the homeostasis and activation of CL-MSCs through Delta-like 1 homolog (Dlk1), which balances MSCs-MTACs number and the lineage differentiation. Dlk1's function on SCs priming and self-renewal depends on its biological forms and its gene expression is under dynamic epigenetic control. Our findings can be validated in clinical samples and applied to accelerate tooth wound healing, providing an intriguing insight of how to direct SCs towards tissue regeneration.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Incisivo/citologia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/metabolismo , Animais , Proteínas de Ligação ao Cálcio/genética , Diferenciação Celular , Linhagem da Célula , Dentina , Epigenômica , Feminino , Expressão Gênica , Homeostase , Humanos , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Knockout , Modelos Animais , Dente Serotino , Ratos , Ratos Wistar , Transdução de Sinais , Nicho de Células-Tronco/fisiologia , Cicatrização
20.
EBioMedicine ; 46: 368-380, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31383551

RESUMO

BACKGROUND: The imprinted gene Delta like non-canonical Notch ligand 1 (Dlk1) is considered an inhibitor of adipogenesis, but its in vivo impact on fat mass indeed remains elusive and controversial. METHODS: Fat deposits were assessed by MRI and DXA scanning in two cohorts of non-diabetic men, whereas glucose disposal rate (GDR) was determined during euglycemic hyperinsulinemic clamp. Blood analyte measurements were used for correlation and mediation analysis to investigate how age, BMI, and fat percentage affect the relation between DLK1 and GDR. Confirmatory animal studies performed in normal (NC) and high fat diet (HFD) fed Dlk1+/+ and Dlk1-/- mice included DXA scanning, glucose tolerance tests (GTTs), blood measurements, and skeletal muscle glucose uptake studies by positron emission tomography (PET), histology, qRT-PCR, and in vitro cell studies. FINDINGS: Overall, DLK1 is positively correlated with fat amounts, which is consistent with a negative linear relationship between DLK1 and GDR. This relationship is not mediated by age, BMI, or fat percentage. In support, DLK1 also correlates positively with HOMA-IR and ADIPO-IR in these humans, but has no linear relationship with the early diabetic inflammation marker MCP-1. In Dlk1-/- mice, the increase in fat percentage and adipocyte size induced by HFD is attenuated, and these animals are protected against insulin resistance. These Dlk1 effects seem independent of gluconeogenesis, but at least partly relies on increased in vivo glucose uptake in skeletal muscles by Dlk1 regulating the major glucose transporter Glut4 in vivo as well as in two independent cell lines. INTERPRETATION: Thus, instead of an adipogenic inhibitor, Dlk1 should be regarded as a factor causally linked to obesity and insulin resistance, and may be used to predict development of type 2 diabetes. FUND: The Danish Diabetes Academy supported by the Novo Nordisk Foundation, The Danish National Research Council (#09-073648), The Lundbeck Foundation, University of Southern Denmark, and Dep. Of Clinical Biochemistry and Pharmacology/Odense University Hospital, the Swedish Research Council, the Swedish Diabetes Foundation, the Strategic Research Program in Diabetes at Karolinska Institute and an EFSD/Lilly grant.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Impressão Genômica , Glucose/metabolismo , Resistência à Insulina , Músculo Esquelético/metabolismo , Obesidade/genética , Obesidade/metabolismo , Tecido Adiposo/metabolismo , Adiposidade , Adulto , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Linhagem Celular , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Obesidade/diagnóstico por imagem , Tomografia Computadorizada com Tomografia por Emissão de Pósitrons , Adulto Jovem
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA